Epidermal growth factor and three-dimensional scaffolds provide conducive environment for differentiation of mouse embryonic stem cells into oocyte-like cells

Epidermal growth factor and three-dimensional scaffolds provide conducive environment for differentiation of mouse embryonic stem cells into oocyte-like cells
azam soleimani,^1,* Soghra Bahmanpour,² Nehleh Zarei Fard,³ Tahereh Talaei-Khozani,⁴
1. Stem Cell Research Laboratory, Department of Anatomical Sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
2. Stem Cell Research Laboratory, Department of Anatomical Sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
3. Stem Cell Research Laboratory, Department of Anatomical Sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
4. Stem Cell Research Laboratory, Department of Anatomical Sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
Introduction: Three-dimensional (3D) culture provides a biomimicry of the naive microenvironment that can support cell proliferation, differentiation and regeneration. Moreover, due to the importance of biomaterials in tissue engineering, successful emulation of tissues depends on the introduction of essential biomolecules, such as growth factors to develop tissue-like constructs. Some growth factors, such as epidermal growth factor (EGF), facilitate normal meiosis during oocyte maturation in vivo. In this study, a scaffold-based 3D co-culture system, using purified alginate was applied to induce oocyte differentiation from mouse embryonic stem cells (mESCs).
Methods: Mouse embryonic stem cells were induced to differentiate into oocyte-like cells using embryoid body (EB) protocol in the two-dimensional (2D) or 3D microenvironment in-vitro. To increase the efficiency of the oocyte-like cells differentiation from mESCs, we employed a co-culture system using ovarian granulosa cells in the presence or absence of epidermal growth factor (+EGF or –EGF) for 14 days and then the cells were assessed for germ cell differentiation, meiotic progression and oocyte maturation markers
Results: The cultures exposed to EGF in the alginate-based 3D microenvironment, showed the highest level of premeiotic (Oct4, Mvh), meiotic (Scp1, Scp3, Stra8, Rec8) and oocyte maturation (Gdf9, Cx37, Zp2) marker genes (P<0.05) in comparison to other groups.
Conclusion: Our data highlight the importance of tissue engineering approach in differentiation of ESCs to oocyte-like cells, and supplementation of EGF is effective in improving the differentiation process. The expression of specific genes related to premeiotic, meiotic and oocyte maturation, accompanying Oct-4, Stra8 and Mvh protein expression confirms the synergistic effect of EGF and alginate-based 3D culture condition in oocyte differentiation.
Keywords: Growth Factor, Alginate, Scaffold, Embryonic Stem Cells, Cell differentiation